Project Summary/Abstract Group 2 innate lymphoid cells (ILC2) are tissue-resident lymphocytes with emerging roles in tissue development, remodeling, and allergic disease. In the lung, ILC2 are key instigators of allergic asthma, yet little is known about where ILC2 reside and how stromal interactions shape ILC2 development and function. We have found that ILC2 in the lung reside in specific perivascular `niches', where they intimately associate with a mesenchymal cell subset that expresses the cytokine IL-33. The goal of this proposal is to rigorously investigate the role and cross-talk of these `niche' mesenchymal cells and their IL-33 production on ILC2 developmental localization, function, and induction of type 2 immune responses. Based on our preliminary data, we propose that niche perivascular mesenchymal cells impact ILC2 localization and function via the production of cytokine signals, including IL-33 and TSLP, providing a nidus for adaptive Th2 immune responses. During inflammation, ILC2 cooperate with additional lymphocytes to cross-regulate mesenchymal cell function and cytokine production. In Aim One, we will define how lung ILC2 `niche-associated' mesenchymal cells and their IL-33 production impact ILC2 activation and induction of adaptive type 2 immune responses. Global deletion of niche mesenchymal cells impaired lung type 2 immune responses, and we predict that cytokines such as IL-33 and TSLP, as well as other unknown factors, contribute to the activation of ILC2 and the type 2 immune response. In Aim Two, we will characterize the stromal cells and signals that govern the postnatal, developing ILC2 niche. IL-33 is known to activate lung ILC2 during development, and our preliminary data indicate the earliest ILC2 interact with mesenchymal niche cells. We will test the development of these perivascular niches and their contribution to ILC2 localization and function in perinatal life. In Aim three, we will explore the cells and cytokines that cross-regulate mesenchymal cells at the ILC2 niche. We have found that both ILC2 and IL-33 expressing mesenchymal niche cells expand during helminth infection of the lung. Here we will test how lymphocyte-derived cytokines impact mesenchymal niche function during both perinatal development and adult immune challenge. We expect that cytokines from both ILC2, as well as recruited lymphocytes, dynamically regulate niche mesenchymal cell function and IL-33 expression to cause long-lasting changes in tissue immune tone and predisposition towards allergic asthma. Together, these three aims use novel techniques to answer a vital and unknown question: how are ILC2 locally regulated at their lung niches? We anticipate these studies will have fundamental implications for the control of ILC2 and type 2 allergic immunity in allergic asthma, and yield insight into how ILC2 impact both beneficial and pathologic type 2 immune responses in the lung and throughout the body.